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- Title
Purification of Paracoccidioides brasiliensis catalase P: subsequent kinetic and stability studies.
- Authors
Chagas, Ronney Fernandes; Bailão, Alexandre Melo; Fernandes, Kátia Flávia; Winters, Michael S.; Pereira, Maristela; Soares, Célia Maria de Almeida
- Abstract
Catalases are essential components of the cellular equipment to cope with oxidative stress. Here we have purified a highly abundant catalase P of Paracoccidioides brasiliensis (PbCatP) that is preferentially expressed in the parasitic yeast phase. This oxidative stress-induced protein was isolated from yeast cells grown in the presence of 15 mM of hydrogen peroxide (H2O2). We have used consecutive steps of protein precipitation and gel filtration chromatography to achieve the purified protein. Protein purification was validated using matrix-assisted laser desorption ionization time-of-flight mass spectrometry and bioinformatics analysis. The purified enzyme showed strong similarity to small-subunit catalases. Like most monofunctional catalases, PbCatP is a homotetramer, resistant to inactivation by acidic conditions, temperature and denaturants. Furthermore, the kinetic behaviour of catalase P was observed to be different at low compared to high H2O2 concentrations. The results demonstrated that a purified PbCatP is a homotetrameric enzyme, classified as a small subunit catalase.
- Subjects
CATALASE; OXIDATIVE stress; PARACOCCIDIOIDES brasiliensis; PROTEINS; HYDROGEN peroxide; GEL permeation chromatography; MATRIX-assisted laser desorption-ionization; MASS spectrometry
- Publication
Journal of Biochemistry, 2010, Vol 147, Issue 3, p345
- ISSN
0021-924X
- Publication type
Article
- DOI
10.1093/jb/mvp182